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Endostatin suppresses growth and progression of many tumors through binding to endothelial cell surface and extracellular matrix proteins like integrin, heparin, matrix metalloproteinase-2 and transglutaminase-2. There is an arginine rich motif on the surface of endostatin that is essential for binding to some of aforementioned proteins. It has been shown that a 27 amino acid peptide derived from amino terminal of endostatin responsible for its anti-angiogenic and anti-tumor activities and mutation of histidines bound to Zn significantly reduce its activity. In the present study, as regards the importance of Zn-binding loop in amino terminal and arginine 27 in carboxyl terminal, peptides corresponding to this region and a mutated variant including isoleusin 26 to arginine mutation synthesized and their structure and interaction with matrix metalloproteinase-2 and transglutaminase-2 analyzed using fluorescence spectroscopy, molecular dynamic and docking simulation techniques. This study aimed to analyze effect of placing two positively charged arginines on the structure and interaction of this fragment of endostatin. Results showed that placing two arginines close together in the carboxyl terminal of peptide increases fluctuations in total structure of peptide, alters Zn-binding loop in the amino terminal and makes binding energy of peptide to matrix metalloproteinase-2 and transglutaminase-2 more negative. It can be inferred that repulsion of two positively charged arginines in carboxyl terminal induces conformational changes in the whole structure and in the amino terminal loop region.

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